In female mammals most oocytes reside in primordial follicles in a resting stage. Little is known about the mechanisms that regulate the movement of these follicles into the growing pool (follicle activation). The pool of resting primordial follicles is a resource, yet untapped, that could be exploited as a source of material to provide alternative methods for alleviating infertility in women and propagating valuable domestic animals and endangered species. It is likely that follicular activation and growth in vivo are regulated by a delicate balance among various factors, both stimulators and inhibitors. To date, efforts to determine the signals that initiate follicle growth have consisted of testing individual "candidates" to determine if they promote or inhibit follicle activation. Although progress has been made using this approach, that progress has been slow. We seek support to develop a complementary approach, based on recent advances in molecular techniques, to determine specific genes that are turned on or off during the activation of follicles. In Specific Aim 1, suppressive subtractive hybridization (SSH) will be used to test the hypothesis that specific genes are turned on (or off) during follicle activation. Candidate genes will be determined by comparing freshly isolated pieces of bovine ovarian cortex (highly enriched for primordial follicles) with cortical pieces cultured for two days (in which 90% of follicles have activated and become primary). Cattle provide an excellent model for human follicular development and SSH are ideally suited for the detection of rare and novel sequences in two closely related cell types/tissues. Specific Aim 2 is designed to test candidate genes identified in Specific Aim 1 for a potential role in follicle activation by using in situ hybridization to localize differentially expressed sequences in freshly isolated vs. cultured bovine cortical pieces. At the end of two years, we expect to have identified a number of candidate genes that will provide important clues to the genetic regulation of follicle activation. The results generated by the proposed experiments will "open the door" to many applications, since it will be the first (to our knowledge) molecular and genetic approach to the regulation of follicle activation. Elucidation of these fundamental mechanisms has practical implications for the development of new contraceptive technologies and alleviation of infertility. [unreadable] [unreadable]